The new england
journal of medicine
n engl j med 375;8 nejm.org August 25, 2016
717
established in 1812
August 25, 2016
vol. 375 no. 8
The authors’ full names, academic de-
grees, and affiliations are listed in the
Appendix. Address reprint requests to
Dr. Cardoso at the Breast Unit, Champali-
maud Clinical Center–Champalimaud
Foundation, Lisbon, Portugal, or at
mindact@ eortc . be.
* A complete list of the Microarray in
Node-Negative and 1 to 3 Positive Lymph
Node Disease May Avoid Chemotherapy
(MINDACT) investigators is provided in
the Supplementary Appendix, available
at NEJM.org.
Drs. Cardoso, van’t Veer, and Bogaerts
and Drs. Rutgers and Piccart contributed
equally to this article.
This article was updated on August 25,
2016, at NEJM.org.
N Engl J Med 2016;375:717-29.
DOI: 10.1056/NEJMoa1602253
Copyright © 2016 Massachusetts Medical Society.
BACKGROUND
The 70-gene signature test (MammaPrint) has been shown to improve prediction of
clinical outcome in women with early-stage breast cancer. We sought to provide prospec-
tive evidence of the clinical utility of the addition of the 70-gene signature to standard
clinical–pathological criteria in selecting patients for adjuvant chemotherapy.
METHODS
In this randomized, phase 3 study, we enrolled 6693 women with early-stage breast cancer
and determined their genomic risk (using the 70-gene signature) and their clinical risk (using
a modified version of Adjuvant! Online). Women at low clinical and genomic risk did not re-
ceive chemotherapy, whereas those at high clinical and genomic risk did receive such therapy.
In patients with discordant risk results, either the genomic risk or the clinical risk was used
to determine the use of chemotherapy. The primary goal was to assess whether, among pa-
tients with high-risk clinical features and a low-risk gene-expression profile who did not re-
ceive chemotherapy, the lower boundary of the 95% confidence interval for the rate of 5-year
survival without distant metastasis would be 92% (i.e., the noninferiority boundary) or higher.
RESULTS
A total of 1550 patients (23.2%) were deemed to be at high clinical risk and low ge-
nomic risk. At 5 years, the rate of survival without distant metastasis in this group was
94.7% (95% confidence interval, 92.5 to 96.2) among those not receiving chemotherapy.
The absolute difference in this survival rate between these patients and those who re-
ceived chemother apy was 1.5 percentage points, with the rate being lower without chemo-
therapy. Similar rates of survival without distant metastasis were reported in the subgroup
of patients who had estrogen-receptor–positive, human epidermal growth factor receptor
2–negative, and either node-negative or node-positive disease.
CONCLUSIONS
Among women with early-stage breast cancer who were at high clinical risk and low
genomic risk for recurrence, the receipt of no chemotherapy on the basis of the 70-gene
signature led to a 5-year rate of survival without distant metastasis that was 1.5 percent-
age points lower than the rate with chemotherapy. Given these findings, approximately
46% of women with breast cancer who are at high clinical risk might not require chemo-
therapy. (Funded by the European Commission Sixth Framework Program and others;
ClinicalTrials.gov number, NCT00433589; EudraCT number, 2005-002625-31.)
abstract
70-Gene Signature as an Aid to Treatment Decisions
in Early-Stage Breast Cancer
F. Cardoso, L.J. van’t Veer, J. Bogaerts, L. Slaets, G. Viale, S. Delaloge, J.-Y. Pierga, E. Brain, S. Causeret,
M. DeLorenzi, A.M. Glas, V. Golfinopoulos, T. Goulioti, S. Knox, E. Matos, B. Meulemans, P.A. Neijenhuis, U. Nitz,
R. Passalacqua, P. Ravdin, I.T. Rubio, M. Saghatchian, T.J. Smilde, C. Sotiriou, L. Stork, C. Straehle, G. Thomas,
A.M. Thompson, J.M. van der Hoeven, P. Vuylsteke, R. Bernards, K. Tryfonidis, E. Rutgers, and M. Piccart,
for the MINDACT Investigators*
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n engl j med 375;8 nejm.org August 25, 2016
718
The
new england journal
of
medicine
W
omen with early-stage breast
cancer are often treated with adjuvant
systemic therapy consisting of chemo-
therapy, endocrine therapy, agents against human
epidermal growth factor receptor 2 (HER2), or
combinations of these drugs when appropriate.
Treatment decisions are based on characteristics
of the tumor (hormonal receptor and HER2 status,
tumor grade and size, and lymph-node status) and
of the patient (age, menopausal status, and per-
formance status).
1
Tools that incorporate these
features, such as Adjuvant! Online
2,3
and PREDICT
Plus,
4
were created to assist in such decision
making. However, these algorithms do not take
into account the individual biologic characteris-
tics of the patient’s tumor.
5
A substantial num-
ber of patients with breast cancer are overtreated
and are thus being exposed to the risk of toxic
effects from adjuvant therapy without deriving
significant benefit. In 2007, respondents to an
international survey identified the use of mo-
lecular signatures to select patients who could
be spared adjuvant therapy as a high priority.
6
Gene-expression profiling studies have distin-
guished at least four molecularly distinct types of
breast cancer.
7,8
Several genomic tests have been
developed to better predict clinical outcome and
to determine whether the addition of adjuvant
chemotherapy to endocrine therapy is worth-
while.
8,9
One such test, the 70-gene signature
10
(MammaPrint), classifies tumors into groups
that are associated with a good prognosis or a
poor prognosis on the basis of the risk of distant
recurrence at 5 years and at 10 years.
11
An inde-
pendent validation study by the TRANSBIG con-
sortium, a network of some 40 partners in 21
countries associated with the Breast Interna-
tional Group (BIG), confirmed that the 70-gene
signature, which has been approved by the Food
and Drug Administration (FDA), is able to distin-
guish patients who are at significant risk for
distant relapse and death from those at low risk.
12
In this international, prospective, randomized,
phase 3 study, called the Microarray in Node-
Negative and 1 to 3 Positive Lymph Node Dis-
ease May Avoid Chemotherapy (EORTC 10041/
BIG 3-04 MINDACT) study, we sought to provide
evidence of the clinical utility
13-15
of the addition
of the 70-gene signature to standard clinical–
pathological criteria in selecting patients for adju-
vant chemotherapy.
16,17
Here, we report 5-year
outcomes and the results of the treatment ran-
domization for groups with discordance in risk.
Methods
Study Patients
From 2007 through 2011, we enrolled patients at
112 institutions in nine European countries. Eli-
gible patients were women between the ages of
18 and 70 years with histologically confirmed
primary invasive breast cancer (stage T1 or T2 or
operable T3). In the initial study design, all the
patients had to have lymph-node–negative dis-
ease, as described in the protocol, available with
the full text of this article at NEJM.org. As of
August 2009, the protocol was revised to allow
the enrollment of women with up to three posi-
tive axillary nodes.
18
The study design called for
following all the patients according to local
standards for at least 10 years; those receiving
endocrine therapy will be followed for a mini-
mum of 15 years.
Written informed consent was obtained from
all the patients. The protocol review committee
of the European Organization for Research and
Treatment of Cancer (EORTC) and the ethics
committee at each participating site approved
the study.
Prognostic Tools
We used the 70-gene signature to determine
genomic risk and Adjuvant! Online (version 8.0
with HER2 status) (www.adjuvantonline.com) to
determine clinical risk (Fig. S7 in the Supple-
mentary Appendix, available at NEJM.org). De-
tails regarding clinical risk assessment accord-
ing to the modified version of Adjuvant! Online
are provided in Table S13 in the Supplementary
Appendix.
A frozen sample of the resected tumor was
shipped on dry ice to Agendia for molecular
diagnostic testing. The quantification of the
tumor-cell percentage was followed by 70-gene
profiling embedded in a whole-transcriptome
array that is cleared for use by the FDA.
19,20
Categorization into Risk Groups
A low clinical risk was defined as the 10-year
probability of breast-cancer–specific survival with-
out systemic therapy of more than 88% among
women with estrogen receptor (ER)–positive
tumors and more than 92% among women with
ER-negative tumors, to account for the 4-percent-
age-point average absolute benefit of adjuvant
endocrine therapy for ER-positive tumors. Pa-
tients with low-risk disease according to both
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n engl j med 375;8 nejm.org August 25, 2016
719
70-Gene Signature in Early Breast Cancer
clinical and genomic results were advised not to
receive adjuvant chemotherapy, whereas patients
who were categorized as having high-risk dis-
ease by both tests were advised to receive chemo-
therapy.
Patients with discordant results (i.e., either
high clinical risk and low genomic risk or low
clinical risk and high genomic risk) were ran-
domly assigned to the chemotherapy group or the
no-chemotherapy group on the basis of either
the clinical result or the genomic result. The
treatment randomization used a minimization
technique that was stratified according to in-
stitution, risk group, hormone-receptor status
(ER-positive or progesterone [PR]-positive vs.
ER-negative and PR-negative), nodal involvement
(yes or no), age (<50 years vs. ≥50 years), HER2
status (HER2-positive vs. HER2-negative vs. un-
known), axillary treatment (sentinel node only vs.
dissection), and type of surgery (mastectomy
vs. breast conservation).
Additional (optional) randomizations were im-
plemented in which patients who were assigned
to receive adjuvant chemotherapy (either random-
ly because of discordant results or because of
high-risk concordance of both tests) could be
randomly assigned to receive an anthracycline-
containing regimen or a docetaxel-plus-capecita-
bine regimen. Similarly, patients with hormone-
receptor–positive breast cancer could undergo
further randomization to a tamoxifen–letrozole
regimen or a letrozole-only regimen. Details re-
garding the various therapy regimens are pro-
vided in the legend for Figure S7 in the Supple-
mentary Appendix.
Protocol Revisions
A change in the RNA-extraction solution that
was used in the calculation of the 70-gene sig-
nature (a change that was not communicated by
the manufacturer) caused a temporary shift in
the risk calculation from May 24, 2009, to Janu-
ary 30, 2010, at which time the issue was discov-
ered and rectified (Table S5 in the Supplemen-
tary Appendix). Because of this shift, 162 patients
who had been identified as being at high ge-
nomic risk were subsequently identified as being
at low genomic risk with the use of the correct
solution (Tables S2 and S4 in the Supplementary
Appendix). The retroactively recalibrated results
were communicated to the independent data and
safety monitoring committee, to all members
of the ethics committees, to the investigators
(who oversaw informing the patients), and to the
TRANSBIG ethics committee. The clinical effect
of this risk revision was that an additional 28
patients received chemotherapy before the re-
sults were corrected, although no patient was
undertreated (Table S4 in the Supplementary
Appendix). For 113 additional patients, the des-
ignations of clinical or genomic risk were cor-
rected after enrollment, owing mainly to incor-
rect reporting of tumor characteristics at the
time of enrollment (Tables S2 and S3 in the
Supplementary Appendix). The actual risk after
the correction of all types of errors is referred to
as the “corrected risk.”
The sample size was modified during the
trial from 6000 patients to 6600 patients because
the proportion of patients who were designated
as being at low clinical and genomic risk was
higher than was initially projected and because
of the need to compensate for the change in
solution used in RNA extraction.
Study End Points
The primary end point was survival without dis-
tant metastasis (event-free rate at 5 years), as
assessed in a time-to-event analysis. Secondary
end points were the proportion of patients who
received chemotherapy according to the clinical
risk as compared with the genomic risk as well
as overall survival and disease-free survival, as
assessed in time-to-event analyses. Survival with-
out distant metastasis was defined as the time
until the first distant metastatic recurrence or
death from any cause. Disease-free survival was
defined as the time until first disease progres-
sion (locoregional, distant relapse, ipsilateral or
contralateral invasive breast cancer, ductal carci-
noma in situ, or an invasive second primary
cancer) or death from any cause. Overall sur-
vival was defined as the time until death from
any cause. Data for patients who had no event at
the cutoff date for the final analysis were cen-
sored at the time of the last disease assessment
for survival without distant metastasis and for
disease-free survival and at the last follow-up
date for overall survival.
Study Oversight
The trial was overseen by the independent data
and safety monitoring committee of the EORTC.
A logistics pilot study validated the real-time
centralized analysis of frozen samples in several
European countries.
19
The MINDACT pilot phase
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n engl j med 375;8 nejm.org August 25, 2016
720
The
new england journal
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(involving the first 800 enrolled patients) con-
firmed the feasibility of the study.
21
Central
pathological review revealed high concordance
between local and central assessments
22,23
(Table
S12 in the Supplementary Appendix).
All randomizations were performed centrally,
initially at the International Drug Development
Institute and, as of 2010, at the EORTC. Genom-
ic profiling was performed by Agendia. The
drugs that were administered during the study
were provided by Novartis, Sanofi-Aventis, and
F. Hoffmann–La Roche, which had no other role
in the study, were not involved in the collection
or analysis of the data, and did not contribute to
the writing of the manuscript. Data collection
and statistical analyses were performed at the
EORTC. The analyses and the final version of the
manuscript were approved by all the authors,
who vouch for the accuracy and completeness of
the data and the adherence of the study to the
protocol.
Statistical Analysis
The primary analysis was designed to test
whether, among patients with high-risk clinical
features and a low-risk gene-expression profile
who did not receive chemotherapy, the lower
boundary of the 95% confidence interval for the
rate of 5-year survival without distant metastasis
would be 92% (i.e., the noninferiority boundary)
or higher, at a one-sided significance level of
0.025. The primary test was to be performed
when two conditions were met: the standard
error for the rate of survival without distant metas-
tasis at 5 years was 0.01 or less and the percent-
age of patients in the primary-test population
with 5-year follow-up was 33% or more. A two-
sided 95% confidence interval for the 5-year rate
of survival without distant metastasis of more
than 92% was considered to indicate statistical
significance. Under these conditions, this test
has 80% power to reject the null hypothesis if
the true 5-year rate of survival without distant
metastasis is 95%.
In addition, three secondary analyses were
planned. In the first analysis, we evaluated the
outcomes in patients in the discordant-risk
groups according to whether they were assigned
to the chemotherapy group or the no-chemo-
therapy group. In the second analysis, we evalu-
ated outcomes in all patients according to
whether the use of chemotherapy had been rec-
ommended by either clinical risk or genomic
risk alone. To have an unbiased estimate for this
analysis, data for patients in the discordant-risk
groups were doubly weighted, because they were
underrepresented by a factor of two in the re-
sulting sample. In the third analysis, we calcu-
lated the percentage of all enrolled patients who
would be assigned to chemotherapy on the basis
of either clinical risk or genomic risk.
In the first two secondary analyses, we report
the results according to the risk category and
treatment assignment at the time of enrollment
(intention-to-treat population). In the third sec-
ondary analysis and in all analyses according to
risk group (namely, those for outcome, patient
characteristics, eligibility, and adherence), we
report the results according to the corrected
risk group.
Exploratory analyses that were defined pro-
spectively in the protocol or in the statistical
analysis plan or that were deemed to be of high
clinical relevance are so described in the text
and in the Supplementary Appendix. All analy-
ses were performed with the use of SAS soft-
ware, version 9.4 (SAS Institute).
Results
Study Patients
The cutoff date for the current analysis was
March 1, 2016; the median follow-up was 5.0
years. Between 2007 and 2011, a total of 11,288
patients underwent screening and 6693 were
enrolled in the study (Fig. S7 in the Supplemen-
tary Appendix). Of the 4595 patients (40.7%)
who underwent screening but were not enrolled,
the main reasons were the unsuitability of tu-
mor material for testing, a decision by the pa-
tient or an investigator not to participate in the
study, or other ineligibility factor (Table S1 in
the Supplementary Appendix).
The patients were divided into four main
groups, according to their clinical and genomic
risk: low clinical risk and low genomic risk,
which included 2745 patients (41.0%); low clini-
cal risk and high genomic risk, which included
592 patients (8.8%); high clinical risk and low
genomic risk, which included 1550 patients
(23.2%); and high clinical risk and high genomic
risk, which included 1806 patients (27.0%).
These numbers were calculated on the basis of
the corrected risk (Fig. 1, and Table S2 in the
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721
70-Gene Signature in Early Breast Cancer
Figure 1. Enrollment and Risk Groups Included in the Analyses.
Patients were categorized into four risk groups on the basis of the 70-gene signature to determine genomic risk and Adjuvant! Online to determine clinical risk. Patients were fur-
ther categorized according to their “corrected risk” after the discovery of risk changes due to factors including a change in the RNA-extraction solution while the study was ongoing
(genomic risk) and incorrect reporting of clinical risk at enrollment (clinical risk). In the analyses according to risk group (namely, those for outcome, patient characteristics, eligibil-
ity, and adherence), the results are reported according to the corrected risk. The primary analysis was conducted in the primary-test population. This population included patients
at high clinical risk and low genomic risk, who were randomly assigned to use the genomic risk for the decision to forgo chemotherapy and who adhered to the treatment assign-
ment of no chemotherapy. Patients with changes in clinical or genomic risk were excluded from the primary-test population. Treatment-randomization analyses for the groups with
discordant clinical and genomic risks were performed with the use of the risk at enrollment (intention-to-treat population). In the intention-to-treat population, patients were ana-
lyzed according to the randomized group, irrespective of adherence. The treatment-randomization analyses were repeated in the per-protocol population, which excluded patients
who were ineligible, had a change in their clinical or genomic risk, or were nonadherent to the treatment assignment. (Patients could have more than one reason for exclusion.)
Per-Protocol Population
Primary-
test
Population
6693 Women with invasive early-stage breast
cancer were enrolled
1873 Had high clinical risk and high
genomic risk at enrollment
1806 Had high clinical risk and high
genomic risk after correction
2634 Had low clinical risk and low
genomic risk at enrollment
2745 Had low clinical risk and low
genomic risk after correction
690 Had low clinical risk and high
genomic risk at enrollment
592 Had low clinical risk and high
genomic risk after correction
1497 Had high clinical risk and low
genomic risk at enrollment
1550 Had high clinical risk and low
genomic risk after correction
690 Underwent randomization 1497 Underwent randomization
4 Were ineligible
53 Had a change
in risk
42 Received chemo-
therapy
4 Were ineligible
57 Had a change
in risk
76 Did not receive
chemotherapy
5 Had unknown
chemotherapy
status
344 Were assigned to
receive chemotherapy
346 Were not assigned
to receive chemotherapy
254 Were included in
the per-protocol
population
224 Were included in
the per-protocol
population
12 Were ineligible
21 Had a change
in risk
85 Received chemo-
therapy
1 Had unknown
chemotherapy
status
11 Were ineligible
26 Had a change
in risk
128 Did not receive
chemotherapy
9 Had unknown
chemotherapy
status
749 Were assigned to
receive chemotherapy
748 Were not assigned
to receive chemotherapy
636 Were included in
the per-protocol
population
21 Had a change
in risk
85 Received chemo-
therapy
1 Had unknown
chemotherapy
status
644 Were included in
the primary-test
population
592 Were included in
the per-protocol
population
Intention-to-Treat Population
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